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209 a.a.
192 a.a.
_MG
Key reference
Full text Protein Sci 12:2622-2632 (2003) PubMed id: 14573873
Structure and stability of the ankyrin domain of the Drosophila Notch receptor. M.E.Zweifel, D.J.Leahy, F.M.Hughson, D.Barrick. ABSTRACT
The Notch receptor contains a conserved ankyrin repeat domain that is required for Notch-mediated signal transduction. The ankyrin domain of Drosophila Notch contains six ankyrin sequence repeats previously identified as closely matching the ankyrin repeat consensus sequence, and a putative seventh C-terminal sequence repeat that exhibits lower similarity to the consensus sequence. To better understand the role of the Notch ankyrin domain in Notch-mediated signaling and to examine how structure is distributed among the seven ankyrin sequence repeats, we have determined the crystal structure of this domain to 2.0 angstroms resolution. The seventh, C-terminal, ankyrin sequence repeat adopts a regular ankyrin fold, but the first, N-terminal ankyrin repeat, which contains a 15-residue insertion, appears to be largely disordered. The structure reveals a substantial interface between ankyrin polypeptides, showing a high degree of shape and charge complementarity, which may be related to homotypic interactions suggested from indirect studies. However, the Notch ankyrin domain remains largely monomeric in solution, demonstrating that this interface alone is not sufficient to promote tight association. Using the structure, we have classified reported mutations within the Notch ankyrin domain that are known to disrupt signaling into those that affect buried residues and those restricted to surface residues. We show that the buried substitutions greatly decrease protein stability, whereas the surface substitutions have only a marginal affect on stability. The surface substitutions are thus likely to interfere with Notch signaling by disrupting specific Notch-effector interactions and map the sites of these interactions. Selected figure(s)
Figure 3.
Figure 3. Superposition of ankyrin repeat structures. (A) Ctrace of a superposition of the three copies of the Notch ankyrin domain present in the asymmetric unit. The superposition was made from repeat two to repeat seven. Chain A is red, chain B is green, and chain C is blue. (B) C
. (C) Superposition of the Notch ankyrin repeat domain (chain A, red) with GABPß (PDB ID 1AWC [PDB] ; purple) and I
B
Figure 4.
Figure 4. Interactions among Notch ankyrin repeat polypeptides within the crystal lattice. (A) Molecular surface representation of the three polypeptides present in the asymmetric unit (chain A, red; chain B, green; and chain C, blue) and chain B' (light green) from the adjacent unit cell. The approximate threefold screw axis runs in the vertical direction on the page. (B) GRASP (Nicholls et al. 1991) potential surface depicting the interface between chains B' (upper right) and A (lower left). The magnesium ion associated with Asp 136 of chain B' and located at the interface between these two chains is colored yellow. (C) Location of interchain salt bridges at the interface between ankyrin chain A (repeats 5-7, gray) and repeats 2-4 of chain B' (repeat 2, orange; repeat 3, yellow; repeat 4, green), oriented as in B. This figure was made using MOLSCRIPT (Kraulis 1991) and Raster3D (Merritt and Bacon 1997).The above figures are reprinted by permission from the Protein Society: Protein Sci (2003, 12, 2622-2632) copyright 2003. Figures were selected by the author.
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